Although many of the effects of luteinizing hormone (LH) in the corpus luteum are thought to be mediated by cAMP, the exact mechanism of action of LH is unknown. It is presumed that the LH-induced events in ovarian tissues mediated by cAMP occur via cAMP-dependent protein kinases. Phosphorylation of proteins is one of the major post-translational mechanisms by which cellular functions are regulated. We have recently described another important messenger system in ovarian tissues which is linked to protein phosphorylation. Receptor-mediated increases in second messengers derived from phosphoinositide metabolism (diacylglycerol and inositol phosphates) lead to the activation of a calcium- and phospholipid-dependent protein kinase (C-kinase). The regulation of phosphoinositide metabolism by LH and other hormones and the presence of C-kinase in the corpus luteum suggest that the control of C-kinase activity is related to the function of the corpus luteum. The specific aims of the proposed research are to purify and further characterize this new protein kinase and evaluate its physiological significance in rat and bovine corpora lutea. The enzyme will be purified with the objectives of 1) determining the specific co-factors required for C-kinase activity, 2) producing monoclonal antibodies to C-kinase for the purpose of detection and quantitation of C-kinase, and 3) determining the substrate specificity of C-kinase. The physiological role for C-kinase will be examined by 1) determining whether or not enzyme activity varies throughout the life span of the corpus luteum and 2) determining if enzyme activity is regulated by LH or other factors known to regulate luteal function. The possible involvement of C-kinase in the regulation of luteal function will be determined by examining its role in 1) gonadotropin binding, 2) receptor phosphorylation, 3) adenylate cyclase activation, and 4) progesterone synthesis. These studies will be carried out by monitoring the phosphorylation of both exogenous and endogenous substrate proteins. The proposed studies on the ovarian C-kinase may provide new insight into the mechanism of LH action and establish a link between hormonally-induced changes in C-kinase activation and corpus luteum function.